DESCRIPTION: Pathogenic microorganisms survive and cause damage within the host by expressing specific gene products called virulence factors. Despite emerging evidences indicating that Bacteroides forsythus (Bf) is a crucial periodontopathic bacterium, the exact mechanisms whereby this microorganism initiates and mediates the pathogenic process are virtually unknown because of the limited information on its virulence factors. Therefore, in order to advance understanding of Bf infection, it is required to identify and characterize unknown virulence factors of Bf. Preliminary data presented in this application indicated that comparative 2-dimensional gel electrophoresis (2-DE) could be used to detect differentially expressed proteins of Bf, and the proteins selectively expressed in vivo were thus discovered. The purpose of this application is to identify and characterize in vivo-expressed proteins of Bf using proteomics study. It is expected that this novel approach, combined with the analysis of the Bf genome sequence, will lead to detection, identification, and characterization of potential virulence factors of Bf. In order to achieve this goal, the following Specific Aims are proposed: 1) to detect differentially expressed proteins of Bf by 2-DE. By comparing the protein profiles of in vitro- and in vivo-grown Bf cells using 2-DE, the proteins selectively expressed in vivo will be detected; 2) to Identify and characterize differentially expressed proteins using mass spectrometry and database search. Identity of in vivo-expressed proteins will be determined by mass spectrometry and search for Bf genome and protein database. This approach will provide a foundation for generating novel hypotheses that can be tested in future studies, in which authentic virulence function of the candidate virulence factors can be validated. The results obtained from these studies will assist to achieve a long-term goal of this laboratory: to gain a better understanding of the pathgogenic of Bf by providing more information on its virulence factors.